Biagi, F. Francisco

Pina, P. Azelia

English

Cadernos Espinosanos (E-Journal)

Rev. Inst. Med. trop. São Paulo 6(3):114-116, maio/junho, 1964 UDC 616.995.121 PRESENCE OF ANTIGENS IN CALCAREOUS CORPUSCLES OF CYSTICERCUS Francisco BIAGI F. e Azelia PINA P. SUMMARY Immunofluorescence studies have been performed with histological sections of Cysticercus cellulosae and anticysticercus immune serum. Positive reactions revealed antigens from this parasite playing a role in the immune response of natural in-fections located in calcareous corpuscles of cysticercus. There are no previous re-cords on this fact and it might be the key to obtaining more specific antigenic extracts of some helminths. INTRODUCTION One of the main problems in immunology of helminthiasis is the large number of com-mon antigens which exist in the parasites, producing cross-reactions between different helminthiasis. Some of these antigenic sub-stances never elicit the production of anti-bodies in natural infections (BIAGI et al.1), because they do not pass from the parasite to the host but are present in the antigenic cxtract obtained from the parasite. On account of both academic interest and the need of serological procedures for the diagnosis of several parenteral helminthiasis (BIAGI et al.2), it is desirable to have purifi-cd antigens giving specific reactions. Labo-ratory fractionation of raw extracts of hel-minths carry the risk of modifying the chemical structure of antigenic substances (KENT 4 ), anatomical dissection of the para-site supplies antigens which also cross-react ( ÜLIVER-GONZALEZ 6 ). Immunofluorescency provide5 an opportunity to localize antigens in microscopic structures of parasites and possibility to isolate physically the micro-5copic structures which contain antigens. It Í3 possible, therefore, to obtain in this way rnore specific antigenic extracts. MATERIAL AND METHODS Living Cysticercus cellulosae were obtained from infected pork. Five micra histological sections of this material were prepared by standard paraffin embedding procedures ( GRADWOHL 3 ) with previous fixation in 10% formalin. Anticysticercus immune serum was obtained from pigs naturally infected with cysticerci. Rabbit anti-pig serum was prepared by one intraperitoneal and three intravenous injections the first 4 days of one week, and one intraperitoneal and three intra-venous injections the first 4 days of the next week, of normal pig serum into rabbits; animais were bled one week after the last injection. Good anti-pig serum was labelled with fluorescein-isothiocyanate (MARSHALL et al. 5 ). ln arder to prove the formation of antibodies against pig serum by the rabbit, the Ouchterlony's agar diffusion technique was performed ( ÜUCHTERLONY 7 ), obtaining in all cases marked precipitation lines in the agar after 24 hours. Histological sections were deparaffined, rehydrated, covered with anticysticercus pig serum and left at 37°C for 30 minutes. They were then washed, covered with fluorescein-Department of Microbio!ogy and Parasitology - School of Medicine - University of México. Apartado Postal No 25788 - México 12, D.F. 114 l:i[;\(';l F .. F . & Plfl'A P., A. - Prcscnce o( .:inugen!-1 in culcart."UUIS (.'1J l.l)USl'ICS o( l'YSlkC1"1"U$. Rct:. Mc1t. /rot). Xuo Í'<ttllo fl: 11-1·116. 196-1. lah(•llc_•tl auli-pig r.:1hl,il ~crum anel ldl <li 37"C íor HO mi nult'~. To c·ou11lt'nstai11. l,o\' illt" til-humin lal..-lll'd will1 rhodamint> was uscd (S~fl'l'II el ai." 1. Fi11ally. t lw sc·t·t ions \vcre wal5he,I. a eovt>n:lidt· wa~ placcd ovpr g1yce• _. ritw, anel oh::-1.•rwcl iu u stnnda rd íluo1'(•sc·c•11c·y ,uic-rm;l'Opt·. Conl ro1 ~lidc·s \\ t"n· pr<·parNI 11:,:i11~ &'ra írom pig1-- willioul tystic:ucosis. Natural íluon•iwcnc·y of Ih<· parasite wa~ dw<:kt•íl in t111sta j11t·,I til'<.·l ions. S lides we-rt· a.lso prepar-t"'<l with onl)' l h l' rhodaminc alhumin cout1• tersla in. RESUJ.:rs ANO C'OM.:'•H:NTS Throughoul this p1•oct'Cl11rt• it \\'Mi ~urpri~• ü,g 10 íin.-1 lhal mo~I of 1h1· íluort•Af•f' in ap• pc~ars in C'alcare-ous (:<>1·pus;c-l<'~. whieh wt>re tomplt·kly §!rt•en wht•11 a11 li<-y~1in•,-r1.1s ~ ra wc-n• us(•d ( fig. 1) a,HI Wt•r<· 11011-rluorl"S<:<'lll in St'clions l re-ah'd wiLh oormal pig $Na ( F'ig. 2). Lcss íluor('S(·c:-in appt•ilrs. if1 1fw nwm-hrane or in o lhf'r pla,·t•:-. o( S4:Úln: oí Ih<- l'ys· ticercus and norw in l lw <-ul ic·lt.·. Rho,fomine-<'Ot111lc_•n;tuin wus lhe~ only onf> prt>sc~n1 in lhe c·111idc•. wos ~l l:-,o dislr ibull'cl in the parasilt.;ti J1Q<I)' and ah::cnl in c.akareous corpuscks. Fig . 1 - Bri~hl c·ul<'an..·ous ('OrPUs<'k-$ (ovaJ 8ll <l!)NI) l'>f!c•ause of green flu Orés.t.•('n ('y l n •• h lNlQloglNII Sef'tion or Cy.,it('CT("ll8 ('(Hllt-/ (Ji'i(l(J trCl.ilNI wlth anli<"ystic:ercus ~nl. Fig. 2 - Oark calcarcous corpu$('les <non• n uorest"ent) ln a sectlon treated wllh non· lmmune !lern. DISCUSSlON Con~idning thal autit·}•sti•·cr(·us s<.·ru from natural i11kc:1ic)11$ Wf•)'(" u::<-d. the presen<.:e o( fluort>:-sc.:cin js io1,•rpre1t>d as •present in an.l i• gpnic suhi;t-8 11<''('$ vthlc.'h plily a rolt> in lhe immunt> rf'sponse du ring a natural if1Íedion; l lic.·n, C'alrare-ous corpusdes nppea r i)::i micro• :-:copie strurlu rt·s having a for~er qmmtity of an1igc~ns. Thetre fimlings \\'l'l'(' tcnfirmccl in !-°:l'Vt'ral instan<'es with iwra from difíNenl pigs. Na.1ural fluon•sc.·c·ncy (gret·n ) wa~ oh-ti(.'íVt·d onl>' i11 thc- hooks. K 11owlc.·dgt· o í c·alc-areous corpm:clcs is meagn·; lhe.·)' diMpJwar during incubat ion io non·llutritivt· ntt'dia~ and mav have a ro le as a huHcr agaitB1 c•xt1·aneo~1s <t<·icls ( VO:'l RnANu 11 ) , Po~ ihly ~)lflt• of tlw p rolt>ins a nd poly:iac.-char ides of lhe_• e.a ln~ rt·oui; cor-1H1srlt>~ leavC' lhe- parasit<· d u ring i 1:,., mt"la• holism Hüd pa.,s to the hosl lo elidi. lhe produrtiorl of antihod ies. Thi~ fio<liog is impor1an1 rn orclf'r to pr(•· JHll'(' lw1t<· 1· .-wti~••nic <'Xlracl~ hf'cause- it i~ very N1~y 10 ::qmn•k cale-a re-ou~ <·orpusclt•i; from olher slru<.·lun·s of lhe pa ra~ilc and in thi5 way the- antigrn ic exlratt will lw lt>~s c·o111ami11a1ed wilh suhslances from otlwr pla(·t•:t or lhe parasite- whid1 cio not pfay a rol1.· io 1lw immurw r1•5=.f)()ll:-l(• i1l l lu• naturul infrrtion. Knowing c·nknn·ou:; corpuscles' 115 BIAGI F., F. & PINA P., A. - Presence of antigens in calcareous corpuscles of cysticercus. Rev. Med. trop. São Paulo 6:114-116, 1964. specific functions, it is possible that their chemical composition will be of less complex substances than the whole parasite. It is possible, therefore, that the number of un-specific antigens will be less in extracts ob-tained from calcareous corpuscles. RESUMO Presença de antígenos nos corpúsculos calcários do cisticerco. Estudos de imunofluorescência foram fei-tos em cortes histológicos de Cysticercus cel-lulosae com sôro imune anticisticerco. As reações positivas revelaram a presença de antígenos dêste parasito, desenvolvendo uma função na resposta imunológica às infecções naturais, localizada nos corpúsculos calcários do cisticerco. Não há registro anterior sô-bre êste fato que pode ser a chave para a obtenção de extratos antigênicos mais espe-cíficos, de alguns helmintos. REFERENCES 1. BIAGI, F., F.; PAYAN, A. M.; CUENCA, G. & HERNANDEZ-CHINAS, J. C. - Value of skin test and precipiting reaction in ctiagnosis of ascariasis. Journal of Infectious Diseases 107: 149-154, 1960. 2. BIAGI, F., F.; PINA, A. & SUARES, A. -Eosinofilia elevada con manifestaciones vis-cerales; III. Estudios serológicos con ex-tractos antígenos de helmintos. Prensa Mé-dica Mexicana 26: 244-248, 1961. 116 3. GRADWOHL, R. B. H. - Clinical Laboratory Methods and Diagnosis. Fifth edition. Mosby Co., 1956. 4. KENT, H. N. - Aspects biochimiques de la specificité chez les Cestoctes. First Sympo-sium of Host Specificity, Institute of Zoology, Neuchatel 293, 1957. 5. MARSHALL, J. D.; EVELAND, W. C. & SMITH, C. W. - Superiority of fluorescein-isothiocyanate (Riggs) for fluorescent anti-body technique with a modification of its application. Proceedings of the Society of Experimental Biology and Medicine 98:898-900, 1958. 6. OLIVER-GONZALEZ, J. - Antigenic analysis of the isola ted tissues and , body fl uids of the roundworms. Journal of Infectioi,s Diseases 72:202-212, 1943. 7. OUCHTERLONY, O. - Diffusion in gel me-thods for immunological analysis. Progress in Allergy 5:1-78, 1958. 8. SMITH, C. W.; MARSHALL, J. D. ,& EVE-LAND, W. C. - Use of contrasting fluores-cent dye as counterstain in fixed tissue pre-parations. Proceedings of the Society of Ex-perimental Bio lo gy and Medicine 102 : 179-181, 1959. 9. von BRAND, T.; MERCADO, T. I.; NYLEN, M. U. & SCOTT, D. B. - Observations on function, composition and structure of cestocte calcareous corpuscles. Experimental Parasi-tology 9:205-214, 1960. Recebido para publicação em 1• novembro 196::l. 

Presence of antigens in calcareous corpuscles of Cysticercus.

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Presence of antigens in calcareous corpuscles of Cysticercus.

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